Cardiac myocytes from embryonic chick and adult rat ventricle will be used to study biochemical responses to muscarinic cholinergic and alpha-adrenergic receptor stimulation. Studies will focus on the hypothesis that phosphatidylinositol turnover is a primary receptor-mediated event that leads to other known biochemical and physiological responses in the myocytes. A role for inositol trisphosphate (IP3) as a second messenger will be suggested by examining the kinetics of formation of all three [3H]inositol phosphates and identifying the isomers of IP3 formed in agonist-treated cardiac cells. A role for diacylglycerol (DAG) as a second messenger will be suggested if agonist stimulation is shown to increase the accumulation of DAG in the cardiac myocyte. The activation of protein kinase C will be assessed by comparing enzyme activity and phorbol ester binding in the cytosol and particulate fractions. Several biochemical responses will be examined as possible consequences of receptor mediated phosphoinositide hydrolysis: increases in cytosolic calcium (as measured by fura-2 fluorescence) or in 45Ca++ fluxes, release of arachidonic acid, formation of cyclic GMP and activation of phosphodiesterase. Physiological events to be examined (in collaboration with other investigators) and correlated with alterations in PI metabolism are increases in cardiac contractility and changes in specific ion conductances in patch-clamped myocytes. Relationships will be established indirectly by comparing agonist and antagonist sensitivities of the PI response with that of other responses and by establishing the effects of pertussis toxin and phorbol esters on these responses. A second messenger role for [IP3] will be examined more directly by determining if exogenous IP3 can mimic the particular response in question or if blockade of IP3 degradation can accentuate the effect of hormone. Similarly, DAG will be increased intracellularly, applied exogenously or its degradation blocked, to mimic or enhance hormonal effects mediated through this putative second messenger.